Overexpression of Thellungiella halophila H(+)-pyrophosphatase Gene Improves Low Phosphate Tolerance in Maize

Low phosphate availability is a major constraint on plant growth and agricultural productivity. Engineering a crop with enhanced low phosphate tolerance by transgenic technique could be one way of alleviating agricultural losses due to phosphate deficiency. In this study, we reported that transgenic maize plants that overexpressed the Thellungiella halophila vacuolar H(+)-pyrophosphatase gene (TsVP) were more tolerant to phosphate deficit stress than the wild type. Under phosphate sufficient conditions, transgenic plants showed more vigorous root growth than the wild type. When phosphate deficit stress was imposed, they also developed more robust root systems than the wild type, this advantage facilitated phosphate uptake, which meant that transgenic plants accumulated more phosphorus. So the growth and development in the transgenic maize plants were not damaged as much as in the wild type plants under phosphate limitation. Overexpression of TsVP increased the expression of genes involved in auxin transport, which indicated that the development of larger root systems in transgenic plants might be due in part to enhanced auxin transport which controls developmental events in plants. Moreover, transgenic plants showed less reproductive development retardation and a higher grain yield per plant than the wild type plants when grown in a low phosphate soil. The phenotypes of transgenic maize plants suggested that the overexpression of TsVP led to larger root systems that allowed transgenic maize plants to take up more phosphate, which led to less injury and better performance than the wild type under phosphate deficiency conditions. This study describes a feasible strategy for improving low phosphate tolerance in maize and reducing agricultural losses caused by phosphate deficit stress.     

Zinc deficiency tolerance in maize is associated with the up‐regulation of Zn transporter genes and antioxidant activities

• Zinc (Zn) is an essential micronutrient for the growth and development of plants. However, Zn deficiency is a common abiotic stress causing yield loss in crop plants. This study elucidates the mechanisms of Zn deficiency tolerance in maize through physiological and molecular techniques.
• Maize lines tolerant (PAC) and sensitive (DAC) to Zn deficiency were examined physiologically and by atomic absorption spectrometry (AAS). Proteins, H₂O₂, SOD, POD, membrane permeability and gene expression (using real‐time PCR) of roots and shoots of both maize lines were assessed.
• Zn deficiency had no significant effect on root parameters compared with control plants in PAC and DAC but showed a substantial reduction in shoot parameters in DAC. AAS showed a significant decrease in Zn concentrations in both roots and shoots of DAC but not PAC under Zn deficiency, implying that Zn deficiency tolerance mechanisms exist in PAC. Consistently, total protein and membrane permeability were significantly reduced in DAC but not PAC in both roots and shoots under Zn deficiency in comparison with Zn‐sufficient plants. Real‐time PCR showed that expression of ZmZIP1, ZmZIP4 and ZmIRT1 transporter genes significantly increased in roots of PAC, but not in DAC due to Zn deficiency compared with controls. The H₂O₂ concentration dramatically increased in roots of DAC but not PAC. Moreover, tolerant PAC showed a significant increase in POD and SOD activity due to Zn deficiency, suggesting that POD‐ and SOD‐mediated antioxidant defence might provide tolerance, at least in part, under Zn deficiency in PAC.
• This study provides an essential background for improving Zn biofortification of maize.

     

Screening tolerance to phosphorus starvation and haplotype analysis using phosphorus uptake 1 (Pup1) QTL linked markers in rice genotypes

Phosphorus (P) deficiency tolerance is a pivotal trait for plant growth and development. Most of the commercial modern cultivars lack this trait and reported it as a very serious problem limiting crop productivity. This trait is advantageous if present in modern high yielding varieties as it increases the yield under the phosphorus-deficient soil conditions. With the importance of phosphorus deficiency tolerance, the present investigation was carried out with an objective to screen for tolerance to phosphorus deficiency using solution culture and phosphorus uptake 1 (Pup1) locus linked markers in 30 diverse rice genotypes. A wide range of varied responses to P deficiency in rice genotypes for all the traits were observed. Root length and enzyme activity showed increased mean performance under the − P condition when compared to + P condition. Medium to high heritability estimates were obtained for most of the traits. Correlation analysis showed that the traits: root P content, fresh shoot weight, dry shoot weight, and shoot length showed highly significant correlations with each other under − P conditions. Based on the hydroponics and molecular screening, three genotypes viz., ADT (R) 48, Improved Pusa Basmati 1 and UPLRI 5 were classified as tolerant for its response to P deficiency as they possessed significant increase in desirable root and shoot traits, increased acid phosphatase enzyme and these genotypes also possessed the Pup1 allele for all the five markers. The selected genotypes may be useful for the exploration of novel genes conferring phosphorus deficiency tolerance and used as donor parents in the breeding programs. Absence of this allele in the rice genotypes viz., drought tolerant (Anna (R) 4) and submergence tolerant (CR 1009 Sub 1) may warrant the development of multiple abiotic stress tolerance cultivars for upland and submergence cropping systems in future rice breeding program.Electronic supplementary materialThe online version of this article (10.1007/s12298-020-00903-1) contains supplementary material, which is available to authorized users.     

玉米耐铝毒基因的分离

以抑制消减杂交（SSH）为手段,以玉米对铝敏感的自交系Mo17和耐铝的自交系TL94B为材料,分别构建它们的正向和反向消减文库,分别筛选获得了124、47、103和64个阳性克隆。对文库的鉴定表明,插入片段分布在0.25-1．0kb之间,阳性克隆率在18％左右。对338个阳性克隆进行测序,得到232种表达序列标签（EST）,其中70．2％的EST可推测其功能。结果表明,玉米的铝离子胁迫反应涉及胁迫因子的信号传导、响应基因的转录表达与调控、物质的合成与运输、细胞结构和功能的改变等。     

A Phosphate Transporter Promoter from Arabidopsis thaliana AtPHT1;4 Gene Drives Preferential Gene Expression in Transgenic Maize Roots Under Phosphorus Starvation

Phosphorus (P) stress responsive genes have been identified and characterized, including the high-affinity phosphate transporter AtPHT1;4 from Arabidopsis thaliana. This gene encodes a membrane protein that is primarily expressed in roots under phosphorus deficiency. A 2.3-kb promoter region from AtPHT1;4 has been fused with the β-glucuronidase (GUS) encoding gene and introduced into maize via biolistic bombardment to evaluate its spatiotemporal activity in a heterologous system. AtPHT1;4::GUS expression is detected preferentially in transgenic maize roots under P deficiency. Further analysis of transgenic plants has also revealed that GUS activity is higher in roots than in leaves by about sixfold. These results demonstrate the ability of AtPHT1;4 promoter to direct expression of the reporter gene in a monocot root system under P stress. This property of AtPHT1;4 promoter makes it useful to engineer maize plants to modify the soil’s rhizosphere and increase efficiency of P acquisition under P stress conditions.     

Relationship between plant phosphorus status and the kinetics of arsenate influx in clones ofdeschampsia cespitosa (L.) beauv. that differ in their tolerance to arsenate

Uptake kinetics of arsenate were determined in arsenate tolerant and non-tolerant clones of the grassDeschampsia cespitosa under differing root phosphorus status to investigate the mechanism controlling the suppression of arsenate influx observed in tolerant clones. Influx was always lower in tolerants compared to non-tolerants. Short term influx of arsenate by the high affinity uptake system in both tolerant clones was relatively insensitive to root phosphorus status. This was in contrast to the literature where the regulation of the phosphate (arsenate) uptake system is normally much more responsive to plant phosphorus status. The low affinity uptake system in both tolerant and non-tolerant clones, unlike the high affinity uptake system, was more closely regulated by root phosphate status and was repressed to a much greater degree under increasing root phosphorus levels than the high affinity system.     

The regulation of root growth in response to phosphorus deficiency mediated by phytohormones in a Tibetan wild barley accession

Low phosphorus (LP) causes a dramatic change of root system architecture in plants, which is possibly mediated by signaling pathways of hormones. In order to understand the regulatory mechanisms of the root development under LP, we examined the potential role of phytohormones in response to LP using three barley genotypes, differing in LP tolerance, namely 2 Tibetan wild barley genotypes XZ99 (LP tolerant) and XZ100 (LP sensitive), and a cultivated barley ZD9 (LP moderately tolerant). The results showed that LP stress caused a number of changes in root development, with XZ99 having less primary root growth inhibition, more lateral root and root hair formation than the other two genotypes. Meanwhile, LP stress also resulted in the dramatic changes in plant hormone contents, with changed extent and pattern differing among the three genotypes. The relative expression of genes responsible for indole acetic acid (IAA) and ethylene synthesis in roots also showed a significant difference among genotypes in both control and LP conditions. It can be concluded that the root system of Tibetan wild barley XZ99 adapts to phosphorus deficiency by changing the signal transduction pathway mediated by auxin, ethylene and cytokinins. However, further studies are needed to elucidate the behaviors of the key genes involved in the hormone-related response.     

Low-P tolerance by maize (Zea mays L.) genotypes: Significance of root growth,and organic acids and acid phosphatase root exudation

We investigated some mechanisms, which allow maize genotypes to adapt to soils which are low in available P. Dry matter production, root/shoot-ratio, root length and root exudation of organic acids and acid phosphatase were investigated in four maize genotypes grown under P-deficient and P-sufficient conditions in sterile hydroponic culture. A low-P tolerant, an acid-tolerant and a low-P susceptible genotype of maize were compared with a Swiss commercial cultivar. The study found increased root development and increased exudation of acid phosphatase under P-deficient conditions in all maize genotypes, except for the Swiss cultivar. Effects on root formation and acid phosphatase were greater for the low-P tolerant than for the low-P susceptible, and the acid soil tolerant genotypes. Organic acid contents in root tissues were increased under P deficiency and related to increased PEPC activity. However, the increase in contents was associated with an increase in exudation for the low-P tolerant genotype only. The low-P susceptible genotype was characterized by high organic acid content in roots and low organic acid exudation. The organic acids content in the phloem exudates of shoots was related to root exudation under different P supply, to the difference between lines in organic acids root content, but not to the low-P tolerance or susceptibility of maize genotypes.     

Meta-analysis and candidate gene mining of low-phosphorus tolerance in maize

Plants with tolerance to low-phosphorus （P） can grow better under Iow-P conditions, and understanding of genetic mechanisms of Iow-P tolerance can not only facilitate identifying relevant genes but also help to develop Iow-P tolerant cultivars. QTL meta-analysis was conducted after a comprehensive review of the reports on O, TL mapping for Iow-P tolerance-related traits in maize. Meta-analysis pro- duced 23 consensus QTL （cQTL）, 17 of which located in similar chromosome regions to those previously reported to influence root traits. Meanwhile, candidate gene mining yielded 215 genes, 22 of which located in the cQTL regions. These 22 genes are homologous to 14 functionally characterized genes that were found to participate in plant Iow-P tolerance, including genes encoding miR399s, Pi transporters and purple acid phosphatases. Four cQTL loci （cQTL2-L cQTLS-3, cQTL6-2, and cQTL10-2） may play important roles for low-P tolerance because each contains more original QTL and has better consistency across previous reports.     

The isolation of genes from the resurrection grass Sporobolus stapfianus which are induced during severe drought stress

A modification of the ‘cold plaque’ screening technique (Hodge et al., Plant Journal1992, 2, 257–260) was used to screen a cDNA library constructed from drought‐stressed leaf tissue of the desiccation tolerant (‘resurrection’) grass Sporobolus stapfianus. This technique allowed a large number of clones representing genes expressed at low abundance to be isolated. An examination of expression profiles revealed that several of these genes are induced in desiccation‐tolerant tissue experiencing severe drought stress. Further characterization indicated that the gene products encoded include an eIF1 protein translation initiation factor and a glycine‐ and proline‐rich protein which have not previously been associated with drought stress. In addition, genes encoding a serine/threonine phosphatase type 2C, a tonoplast‐intrinsic protein (TIP) and an early light‐inducible protein (ELIP) were isolated. A number of these genes are expressed differentially in desiccation‐tolerant and desiccation‐sensitive tissues, suggesting that they may be associated with the desiccation tolerance response of S. stapfianus. The results indicate that there may be unique gene regulation processes occurring during induction of desiccation tolerance in resurrection plants which allow different drought‐responsive genes to be selectively expressed at successive levels of water loss.     

Genes mapping to boron tolerance QTL in barley identified by suppression subtractive hybridization

Boron tolerance is a quantitative trait controlled by multiple genes. Suppression subtractive hybridization was carried out on root cDNA from bulked boron tolerant and intolerant doubled haploid barley lines grown under moderate boron stress to identify genes associated with boron tolerance. One hundred and eleven clones representing known proteins were found to be up‐regulated in the tolerant bulk upon boron stress. Nine clones were genetically mapped to previously reported boron tolerance QTL. These include a clone identical to the boron transporter gene Bot1 and a clone coding for a bromo‐adjacent homology domain‐containing protein, mapping to the 6H boron tolerance locus and co‐segregating with reduced boron intake in a Clipper × Sahara‐3771 mapping population. A third clone mapping to the 2H QTL region encoding an S‐adenosylmethionine decarboxylase precursor was found to provide tolerance to high boron by heterologous expression. Yeast cells expressing Sahara SAMDC were able to grow on 15 mm boron solid media and maintained cellular boron concentrations at 13% lower than control cells expressing empty vector. The data suggest that an antioxidative response mechanism involving polyamines and the ascorbate–glutathione pathway in Sahara barley may provide an advantage in tolerating high soil concentrations of boron.     

Effect of plant growth promoting <Emphasis Type="Italic">Pseudomonas</Emphasis> spp. on compatible solutes,antioxidant status and plant growth of maize under drought stress

Drought is one of the major abiotic stresses affecting yield of dryland crops. Rhizobacterial populations of stressed soils are adapted and tolerant to stress and can be screened for isolation of efficient stress adaptive/tolerant, plant growth promoting rhizobacterial (PGPR) strains that can be used as inoculants for crops grown in stressed ecosystems. The effect of inoculation of five drought tolerant plant growth promoting Pseudomonas spp. strains namely P. entomophila strain BV-P13, P. stutzeri strain GRFHAP-P14, P. putida strain GAP-P45, P. syringae strain GRFHYTP52, and P. monteilli strain WAPP53 on growth, osmoregulation and antioxidant status of maize seedlings under drought stress conditions was investigated. Drought stress induced by withholding irrigation had drastic effects on growth of maize seedlings. However seed bacterization of maize with Pseudomonas spp. strains improved plant biomass, relative water content, leaf water potential, root adhering soil/root tissue ratio, aggregate stability and mean weight diameter and decreased leaf water loss. The inoculated plants showed higher levels of proline, sugars, free amino acids under drought stress. However protein and starch content was reduced under drought stress conditions. Inoculation decreased electrolyte leakage compared to uninoculated seedlings under drought stress. As compared to uninoculated seedlings, inoculated seedlings showed significantly lower activities of antioxidant enzymes, ascorbate peroxidase (APX), catalase (CAT), glutathione peroxidase (GPX) under drought stress, indicating that inoculated seedlings felt less stress as compared to uninoculated seedlings. The strain GAP-P45 was found to be the best in terms of influencing growth and biochemical and physiological status of the seedlings under drought stress. The study reports the potential of rhizobacteria in alleviating drought stress effects in maize.